Laboratory capacity expansion: lessons from establishing molecular testing in regional referral laboratories in Ethiopia.

Respiratory viruses contribute to high morbidity and mortality in Africa. In 2020, the Ohio State University's Global One Health Initiative, in collaboration with the Ethiopian Public Health Institute and the US Centers for Disease Control and Prevention, took action to strengthen Ethiopia's existing respiratory virus surveillance system through decentralization of laboratory testing and scale-up of national and regional capacity for detecting respiratory viruses. In August 2022, four regional laboratories were established, thereby raising the number of reference laboratories conducting respiratory virus surveillance to five. This article highlights lessons learned during implementation and outlines processes undertaken for laboratory scale-up and decentralization.

Pre-Harvest Food Safety Challenges in Food-Animal Production in Low- and Middle-Income Countries.

Food safety remains a significant global public health concern, with the risk of unsafe food varying worldwide. The economies of several low- and middle-income countries (LMICs) heavily rely on livestock, posing a challenge to ensuring the production of safe food. This review discusses our understanding of pre-harvest critical issues related to food safety in LMICs, specifically focusing on animal-derived food. In LMICs, food safety regulations are weak and inadequately enforced, primarily concentrating on the formal market despite a substantial portion of the food sector being dominated by informal markets. Key critical issues at the farm level include animal health, a low level of good agriculture practices, and the misuse of antimicrobials. Effectively addressing foodborne diseases requires a comprehensive One Health framework. Unfortunately, the application of the One Health approach to tackle food safety issues is notably limited in LMICs. In conclusion, considering that most animal-source foods from LMICs are marketed through informal channels, food safety legislation and policies need to account for this context. Interventions aimed at reducing foodborne bacterial pathogens at the farm level should be scalable, and there should be strong advocacy for the proper implementation of pre-harvest interventions through a One Health approach.

Antimicrobial Resistance and Virulence Gene Profile of Clinical Staphylococcus aureus: A Multi-Center Study from Ethiopia.

Background: Staphylococcus aureus causes a wide range of infections from mild skin and soft tissue to severe life-threatening bacteremia. The pathogenicity of S. aureus infections is related to various bacterial surface components and extracellular proteins such as toxic-shock syndrome (TSS) toxin and Panton-Valentine leukocidin (PVL). In this study we determine the antimicrobial resistance of isolated strains and their virulence genes in Ethiopia. Methods: A total of 190 archived S. aureus isolates from four Ethiopia Antimicrobial Resistance (AMR) Surveillance sites were analyzed. The identification of S. aureus was done by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF Biotyper) and antimicrobial susceptibility test (AST) was done using VITEK® 2. Multiplex PCR was used to detect mecA, mecC, pvl and spa genes and super-antigens (sea, seb, sec, seh and sej staphylococcal enterotoxins). Results: A total of 172 isolates were confirmed as S. aureus, 9 (5.23%) were methicillin-resistant S. aureus (MRSA) and 163 (94.76%) were methicillin-susceptible S. aureus (MSSA). AST showed that 152 (88.4%) isolates were resistant to penicillin; 90 (52.32%) resistant to trimethoprim-sulfamethoxazole; and 45 (26.16%) resistant to tetracycline. A total of 66 (38.37%) isolates harbored at least one staphylococcal enterotoxin gene and 31 (46.96%) isolates had more than one. The most frequent enterotoxin gene encountered was seb 28 (16.28%). The TSST-1 gene was detected in 23 (13.37%). Presence of staphylococcal enterotoxin gene showed significant association with antibiotic resistance to cefoxitin, benzylpenicillin, oxacillin, erythromycin, clindamycin, tetracycline and SXT. The pvl gene was detected in 102 (59.3%) of isolates. Isolates from patients below 15 years of age showed significantly high numbers of pvl gene (P = 0.02). Presence of sej (P = 0.011) and TSST-1 (P <0.001) genes were associated with the presence of pvl gene. Conclusion: In this study, isolates were highly resistant to oral antibiotics and the pvl, seb, sea and TSST-1 genes were prevalent.

Clonal diversity of Staphylococcus aureus isolates in clinical specimens from selected health facilities in Ethiopia.

Staphylococcus aureus is among the top three causative agents of nosocomial infection in Ethiopia. The majority of studies in Ethiopia have focused on the epidemiology of S. aureus in hospital settings, with limited molecular genotyping results. Molecular characterization of S. aureus is essential for identification of strains, and contributes to the control and prevention of S. aureus infection. The aim of the current study was to determine the molecular epidemiology of methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) isolates recovered from clinical specimens in Ethiopia. A total of 161 MSSA and 9 MRSA isolates were characterized using pulsed-field gel electrophoresis (PFGE) and staphylococcal protein A (spa) typing. Based on the PFGE analysis, MSSA isolates were grouped into eight pulso-types groups (from A to I), while MRSA isolates clustered into three (A, B and C) pulso-types with more than 80% similarity. The spa typing analysis showed diversity of S. aureus with 56 distinct spa types. Spa type t355 was most prevalent (56/170, 32.9%), while eleven new spa types were detected including t20038, t20039, and t20042. The identified spa types were clustered into 15 spa-clonal complexes (spa-CCs) using BURP analysis; novel/unknown spa types were further subjected to MLST analysis. The majority of isolates belonged to spa-CC 152 (62/170, 36.4%), followed by spa-CC 121 (19/170, 11.2%), and spa-CC 005 (18 /170, 10.6%). Of the nine MRSA isolates, 2 (22.2%) were spa-CC 239 with staphylococcal cassette chromosome (SCC)mec III. These findings highlight the diversity of S. aureus strains in Ethiopia, as well as the presence of potentially epidemic strains circulating in the country necessitating further characterization of S. aureus for antimicrobial resistance detection and infection prevention purposes.

Characterization of plasmids carrying blaCTX-M genes among extra-intestinal Escherichia coli clinical isolates in Ethiopia.

CTX-Ms are encoded by blaCTX-M genes and are widely distributed extended-spectrum ß-lactamases (ESBLs). They are the most important antimicrobial resistance (AMR) mechanism to ß-lactam antibiotics in the Enterobacteriaceae. However, the role of transmissible AMR plasmids in the dissemination of blaCTX-M genes has scarcely been studied in Africa where the burden of AMR is high and rapidly spreading. In this study, AMR plasmid transmissibility, replicon types and addiction systems were analysed in CTX-M-producing Escherichia coli clinical isolates in Ethiopia with a goal to provide molecular insight into mechanisms underlying such high prevalence and rapid dissemination. Of 100 CTX-Ms-producing isolates obtained from urine (84), pus (10) and blood (6) from four geographically distinct healthcare settings, 75% carried transmissible plasmids encoding for CTX-Ms, with CTX-M-15 being predominant (n = 51). Single IncF plasmids with the combination of F-FIA-FIB (n = 17) carried the bulk of blaCTX-M-15 genes. In addition, IncF plasmids were associated with multiple addiction systems, ISEcp1 and various resistance phenotypes for non-cephalosporin antibiotics. Moreover, IncF plasmid carriage is associated with the international pandemic E. coli ST131 lineage. Furthermore, several CTX-M encoding plasmids were associated with serum survival of the strains, but less so with biofilm formation. Hence, both horizontal gene transfer and clonal expansion may contribute to the rapid and widespread distribution of blaCTX-M genes among E. coli populations in Ethiopian clinical settings. This information is relevant for local epidemiology and surveillance, but also for global understanding of the successful dissemination of AMR gene carrying plasmids.

Antimicrobial Resistance Profiling and Molecular Epidemiological Analysis of Extended Spectrum ß-Lactamases Produced by Extraintestinal Invasive Escherichia coli Isolates From Ethiopia: The Presence of International High-Risk Clones ST131 and ST410 Revealed.

The treatment of invasive Escherichia coli infections is a challenge because of the emergence and rapid spread of multidrug resistant strains. Particular problems are those strains that produce extended spectrum ß-lactamases (ESBL's). Although the global characterization of these enzymes is advanced, knowledge of their molecular basis among clinical E. coli isolates in Ethiopia is extremely limited. This study intends to address this knowledge gap. The study combines antimicrobial resistance profiling and molecular epidemiology of ESBL genes among 204 E. coli clinical isolates collected from patient urine, blood, and pus at four geographically distinct health facilities in Ethiopia. All isolates exhibited multidrug resistance, with extensive resistance to ampicillin and first to fourth line generation cephalosporins and sulfamethoxazole-trimethoprim and ciprofloxacin. Extended spectrum ß-lactamase genes were detected in 189 strains, and all but one were positive for CTX-Ms ß-lactamases. Genes encoding for the group-1 CTX-Ms enzymes were most prolific, and CTX-M-15 was the most common ESBL identified. Group-9 CTX-Ms including CTX-M-14 and CTX-27 were detected only in 12 isolates and SHV ESBL types were identified in just 8 isolates. Bacterial typing revealed a high amount of strains associated with the B2 phylogenetic group. Crucially, the international high risk clones ST131 and ST410 were among the sequence types identified. This first time study revealed a high prevalence of CTX-M type ESBL's circulating among E. coli clinical isolates in Ethiopia. Critically, they are associated with multidrug resistance phenotypes and high-risk clones first characterized in other parts of the world.

Enterotoxin gene profile of Staphylococcus aureus isolates recovered from bovine milk produced in central Ethiopia.

INTRODUCTION: Staphylococcal food intoxication is dependent on the production of enterotoxins, the single most important virulence factors. Various studies conducted in Ethiopia have depicted the prevalence of S. aureus in bovine milk. However, there is no published data regarding the enterotoxin gene profile of S. aureus isolates in Ethiopia. The aim of this study was, therefore, to evaluate enterotoxin gene carriage profile of S. aureus isolates recovered from bovine milk samples from central Ethiopia. METHODOLOGY: In this study, 109 S. aureus isolates recovered from bovine milk were analyzed for carriage of the classical enterotoxin genes. Genomic DNA extraction was performed using a commercially available kit. Two sets of multiplex polymerase chain reaction (PCR) assays were used to detect the five classical enterotoxin-coding genes and the toxic shock syndrome toxin gene. RESULTS: At least one type of S. aureus enterotoxin gene (SE) was carried in 73 (66.9%) of the isolates. The most frequently encountered gene was sea (40; 36.7%) followed by seb (19; 17.4%), see (18; 16.5%), tst (16; 14.7%), sec-1 (12; 11.01%), and sed (7; 6.4%). Of the 73 S. aureus isolates harboring at least one of the enterotoxin genes, 26 (35.6%) strains harbored more than one enterotoxin gene. CONCLUSIONS: More than half of the S. aureus isolates harbored at least one of the enterotoxin coding genes, indicating milk specimens contaminated by S. aureus could have a high chance of causing food intoxication.

Prevalence of Listeria monocytogenes in raw bovine milk and milk products from central highlands of Ethiopia.

INTRODUCTION: Listeria monocytogenes is of major significance in human and veterinary medicine. Most human Listeria infections are foodborne and the association of contaminated milk and dairy produce consumption with human listeriosis is noteworthy. In Ethiopia, there is limited data regarding the prevalence of L. monocytogenes in raw bovine milk and dairy products. The aim of this study was, therefore, to determine the prevalence of L. monocytogenes in raw bovine milk and dairy produce. METHODOLOGY: A total of 443 milk and milk product samples were microbiologically analyzed following methods recommended by the U.S. Food and Drug Administration Bacteriological Analytical Manual to isolate Listeria spp. RESULTS: The overall prevalence of Listeria spp. was 28.4% and specifically that of L. monocytogenes was 5.6%. Taking the prevalence of Listeria spp. into consideration, cheese was found to be highly contaminated at 60%, followed by pasteurized milk samples (40%), raw milk (18.9%) and yoghurt (5%). Considering the prevalence of Listeria monocytogenes only, raw milk had the lowest contamination while cheese had the highest, followed by pasteurized milk and yoghurt. CONCLUSIONS: Raw milk and milk products produced in urban and peri-urban areas of central Ethiopia were contaminated with pathogenic bacteria, L. monocytogenes. The detection of this pathogen in raw milk and milk products warrants an urgent regulatory mechanism to be put in place and also the potential role of milk processing plants in the contamination of dairy products should be investigated.